Increased activity of the Vesicular Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor TI-VAMP/VAMP7 by Tyrosine Phosphorylation in the Longin Domain

• Published in: The Journal of biological chemistry Vol. 288; no. 17; pp. 11960 - 11972
• Main Authors: Burgo, Andrea, Casano, Alessandra M., Kuster, Aurelia, Arold, Stefan T., Wang, Guan, Nola, Sébastien, Verraes, Agathe, Dingli, Florent, Loew, Damarys, Galli, Thierry
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 26.04.2013; American Society for Biochemistry and Molecular Biology
• Subjects: Animals; Biochemistry, Molecular Biology; Cell Biology; Cellular Biology; Cercopithecus aethiops; Chlorocebus aethiops; COS Cells; CSK Tyrosine-Protein Kinase; Exocytosis; Exocytosis - drug effects; Exocytosis - physiology; HeLa Cells; Humans; Hypoglycemic Agents; Hypoglycemic Agents - pharmacology; Insulin; Insulin - pharmacology; Insulin-Like Growth Factor I; Insulin-Like Growth Factor I - pharmacology; Life Sciences; Membrane Trafficking; Molecular biology; Phosphorylation; Phosphorylation - physiology; Phosphotyrosine; Protein Structure, Tertiary; R-SNARE Proteins - genetics; R-SNARE Proteins - metabolism; SNARE Proteins; SNARE Proteins - genetics; SNARE Proteins - metabolism; Src; src-Family Kinases; src-Family Kinases - genetics; src-Family Kinases - metabolism; Subcellular Processes; TI-VAMP/VAMP7; Membrane Trafficking; TI-VAMP/VAMP7; Exocytosis; Src; SNARE Proteins; Phosphotyrosine
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.415075

Vesicular (v)- and target (t)-SNAREs play essential roles in intracellular membrane fusion through the formation of cytoplasmic α-helical bundles. Several v-SNAREs have a Longin N-terminal extension that, by promoting a closed conformation, plays an autoinhibitory function and decreases SNARE complex formation and membrane fusion efficiency. The molecular mechanism leading to Longin v-SNARE activation is largely unknown. Here we find that exocytosis mediated by the Longin v-SNARE TI-VAMP/VAMP7 is activated by tonic treatment with insulin and insulin-like growth factor-1 but not by depolarization and intracellular calcium rise. In search of a potential downstream mechanism, we found that TI-VAMP is phosphorylated in vitro by c-Src kinase on tyrosine 45 of the Longin domain. Accordingly, a mutation of tyrosine 45 into glutamate, but not phenylalanine, activates both t-SNARE binding and exocytosis. Activation of TI-VAMP-mediated exocytosis thus relies on tyrosine phosphorylation. Background: The mechanism of activation of Longin vesicular SNAREs, particularly TI-VAMP/VAMP7, which is autoinhibited by its Longin domain, is largely unknown. Results: Mimicking tyrosine 45 phosphorylation activates both t-SNARE binding and exocytosis of TI-VAMP. Conclusion: TI-VAMP is positively regulated by tyrosine phosphorylation of the Longin domain. Significance: Exocytosis mediated by TI-VAMP can be regulated by release of Longin domain autoinhibition.